We investigated the antiproliferative effects of the cytoplasmic fraction of Lactococcus lactis ssp. lactis (L.lac CF) on the SNU-1 human stomach cancer cell line. The proliferation of SNU-1 cells was inhibited by treatment with L.lac CF in a time- and dose-dependent manner. L.lac CF caused G0/G1 cell cycle arrest, which was associated with an increase in p53 and p21 expression, the reduction of cyclin D1 expression, and retinoblastoma protein phosphorylation. L.lac CF induced apoptosis in SNU-1 cells, as demonstrated by increased nucleus condensation and a sub-G1 peak. Caspase-3 activation, the induction of p53, and the downregulation of Bcl-2 were also observed in L.lac CF-treated cells. Thus, the inhibitory effect of L.lac CF on SNU-1 cell growth is mainly attributable to the induction of G0/G1 cell cycle arrest and apoptosis.